Apparatus for separation of cryoprecipitate from blood plasma and method

ABSTRACT

A blood collection system which comprises several blood compatible sealed bags connected together by blood compatible conduits. Means for separating cryoprecipitate from blood plasma are provided. This separation is made by a filtering means. The filtering means is positioned in the outlet of one of the bags of the system. After the blood cells have been removed by a prior step, the remaining blood plasma is collected in the bag which has the filtering means in the outlet. In this bag, the blood plasma is frozen and then slowly thawed. Factor VIII rich cryoprecipitate forms in this bag as part of the freezing and thawing process. As the plasma melts, it flows from the bag through the filtering means. The filtering means positioned in the outlet of the bag retains the cryoprecipitate. Thus, the separation of Factor VIII rich cryoprecipitate from the blood plasma is effected.

BACKGROUND OF THE INVENTION

This application relates to an improved blood collection apparatus andmethod in which the separation of Factor VIII rich cryoprecipitate fromthe remainder of the blood plasma is facilitated in an improved mannerand in which the yield of Factor VIII rich cryoprecipitate is improved.

Many thousands of units of blood are collected each year in multiple bagblood collection systems comprising several blood compatible, sealedbags connected together with blood compatible tubing.

In a typical operation, a donor needle is inserted into the vein of apatient. This needle is connected by suitable tubing, such as vinyltubing, to a first blood bag containing a small amount of conventionalblood cell preservative, such as ACD or CPD. The blood is allowed tofill the first blood bag of the system, the donor needle is withdrawnfrom the patient, and the tubing connecting the needle to the bag issealed. Following this, the blood collection system is centrifuged tocause the blood cells to settle and separate from the plasma. The plasmais then expressed through another tubing into a second blood bag, whilethe cells remain in the first blood bag.

Optionally, platelets may be harvested at this stage by a secondcentrifugation.

The plasma in the second blood bag is then frozen, either byrefrigeration or by immersion in a mixture of dry ice and ethanol or asimilar solvent.

After this, the frozen plasma is conventionally allowed to thaw slowlyand then is centrifuged once again to settle solid material in the cold,thawed plasma. This solid material is known as Factor VIII richcryoprecipitate.

After the conventional centrifuging, the plasma, which is nowcryoprecipitate-poor, is expressed through tubing into a third blood bagfor use, leaving behind the Factor VIII rich cryoprecipitate. ThisFactor VIII rich cryoprecipitate is the source of an importanttherapeutic agent for arresting the symptoms of a common type ofhemophilia.

In accordance with this invention, an apparatus and a method of usingthe same is provided which permits the collection of increased yields ofFactor VIII rich cryoprecipitate without the use of a secondcentrifuging step and which provides substantial savings in time andeffort when compared with the present technique for obtaining FactorVIII rich cryoprecipitate.

DESCRIPTION OF THE INVENTION

This improved blood collection system incorporates a plurality of bloodcompatible, sealed bags connected together with blood compatible conduitmeans. Blood collection means, such as a phlebotomy needle connected tovinyl plastic tubing, communicates with the interior of the first of thebags. Accordingly, the blood is collected into the first bag andcentrifuged to remove blood cells. The plasma is then expressed throughthe appropriate connecting conduit from the first bag to the second bag.Optionally, platelets may be harvested by another centrifugation inwhich the platelets remain in the second bag and the plasma is passed onto an additional bag. The bag containing the plasma is sealed, typicallyby heat sealing or clamping of the conduit, and frozen to form theFactor VIII rich cryoprecipitate.

For the final separation step, it is preferred to allow the frozenplasma to thaw slowly, permitting the thawed plasma to pass through thefiltering means and out of the bag into another storage container as itmelts. The thawing process typically can be performed between 2° and 20°C. (preferably between 2°-5° C.) for maximum yield of cryoprecipitate.The frozen bag is hung up and allowed to slowly melt and drain throughthe filtering means and bag outlet into another container. The thawingand draining process takes approximately 18 to 24 hours at 5° C. in aFenwal blood bag containing one unit of blood plasma.

After the drainage is completed, the bag containing the Factor VIII richcryoprecipitate can be sealed and separated from any remaining attachedbags. One unit of cryoprecipitate is obtained (often with an increasedyield of Factor VIII). Likewise, one unit of cryoprecipitate-poor plasmapasses into the storage receptacle, which is usually the third or fourthbag of the system.

The filtering means used in the separation of the cryoprecipitate fromthe plasma must exhibit certain characteristics. It must pass thecryoprecipitate-poor plasma while retaining the Factor VIII richcryoprecipitate. Cryoprecipitate-poor plasma can pass through aconventional screen type or membrane filter with a 2 micron pore.However, the cryoprecipitate clogs present-day screen or membranefilters. Therefore, part of the multiple blood bag system must accountfor and solve the clogging problem. Accordingly, "filtering means" asused herein means a filter which can pass the cryoprecipitate-poorplasma while retaining the Factor VIII rich cryoprecipitate and whichsolves the clogging problem.

Presently, the filtering means most effective in exhibiting thesenon-clogging properties is a depth filter. "A depth filter consists offibrous, granular or sintered materials, pressed, wound, fired, orotherwise bonded into a tortuous maze of flow channels." (Millipore'sHigh Volume Pharmaceutical and Biological Filtration, (1972) page 2.)

Because a depth filter is a three-dimensional irregular maze ofmaterial, it has a large internal surface area on which cryoprecipitatecan be caught. Therefore, it will not readily become clogged as plasmais being passed through the filter.

Because a depth filter is a three dimensional maze of material, itsfiltering capacity cannot be described in terms of the size of theparticles it will pass. Rather, a depth filter's filtering capacity isdefined experimentally in terms of the percentage of particles of acertain size which will pass through it. It follows that outside factorswill affect the filtering rate of depth filters. Flow rate, pressure,and particle adhesiveness are three such factors.

The presently preferred filtering means used in the improved bloodcollection system of this invention is a polyurethane open cell foamdepth filter.

The specific embodiment described below illustrates one exemplary meansfor embodying the invention of this application. A "triple" bag systemis shown bearing some similarity to the currently available Fenwaltriple bags, but other structures such as single, double, and quadruplebag systems may be used if desired, modified in accordance with thisinvention.

In the drawings:

FIG. 1 is a plan view of a triple bag blood collection system, utilizingthe invention of this application, with a portion broken away forshowing construction detail.

FIG. 2 is a sectional view taken along line 2--2 of FIG. 1.

Referring to the drawings, a blood collection system is shown comprisingblood compatible, sealed blood bags 10, 12, 14, connected together withblood compatible conduit means 16, 18, which are shown to be vinylplastic tubes. As stated above, many of the overall specific details ofdesign of bags 10, 12, 14, and their respective conduits 16, 18, arewell known, and similar bags made of vinyl plastic are commerciallyavailable at the present time.

A blood collection needle 20, conventionally sheathed with a needleprotection cover 22, is shown to be connected in conventional manner byvinyl plastic tubing 24 to the interior of the first bag 10.

Vinyl plastic tubing 16 and flexible connector tube 17 providecommunication between first bag 10 and second bag 12, while vinylplastic tubing 18 and similar connector tube 19 provide connectionbetween second bag 12 and third bag 14. Each of the three bags carries apair of outlet ports 26 which are surrounded with conventionalrupturable sheaths 28, specifically shown to be the design presentlyutilized in the Fenwal blood bags.

In accordance with this invention, filtering means is located in theinterior of second bag 12, being positioned to cover outlet port tube 32of bag 12. Outlet port tube 32, which is generally made of plastic, inturn communicates through tube 18 with the interior of bag 14.

Filtering means 30 may be polyurethane open cell foam depth filter soldunder the brand name Scott Felt by the Scott Paper Company.

This material is a thick, sheet-like material, the grade specificallyused herein being approximately 3/8 inch thick. The material, asoriginally made, has approximately 100 pores per inch, but then iscompressed to 1/8th of its thickness, and heat set so that the materialdoes not re-expand significantly after release from the compression.

Referring also to FIG. 2, a slit 34 is shown to be cut longitudinally infiltering means 30. Port tube 32 is positioned within the slit. Porttube 32 and filtering means 30 are positioned as shown in FIG. 1 betweenthe pair of plastic sheets 35, 37 that makes up blood bag 12, and theperiphery 36 of sheets 35, 37 is heat sealed together, with the bottom38 of filtering means 30 between them, so that periphery 36, bottom 38of filtering means 30, and outlet port tube 32 all are sealed togetherinto a unitary, sealed mass. Both ends of outlet port tube 32 protrudefrom opposite sides of the heat sealed periphery 36, for fluidcommunication therethrough.

Tube 19 is connected to port tube 32 by means of sleeve 40, which may bemade of plastic or the like. Sleeve 40 defines a diaphragm 42 whichblocks fluid flow out of bag 12 through port tube 32 intil the diaphragmis ruptured by pointed tubular cannula 44, which is constructed in amanner similar to the diaphragm rupturing system of U.S. Pat. No.3,110,308, and is retained in a bore of connector tube 19, since thebore of tube 18 is preferably too small to receive the cannula.Alternatively, the system for rupturing diaphragm 42 can be similar toU.S. Pat. No. 3,685,795, or any other conventional diaphragm rupturingsystem may be used.

A similar sleeve 40, having diaphragm 42, communicates with the interiorof first blood bag 10, being retained with the corresponding heat sealedperiphery 36 of bag 10 for leak free communication with the interior ofthe bag. Another hollow cannula 44, retained in connector tube 17, ispresent for the same function of opening diaphragm 42.

Sealed tubes 46 on bags 12 and 14 are ports used in the sterilization ofthe bags, to provide venting of the bags during and after thesterilization process. Thereafter the tubes are sealed as shown.

Identical serial numbers may be placed on vinyl tubing 16, 18 and 24 inthe manner of Bellamy U.S. Pat. No. 2,896,619 for identification of therespective bags after they have been separated from each other, and foranalysis of small samples of the contents of the bags.

Hangers 48 at the bottom of bags 10 and 14 permit inversion of the bagson an I.V. pole for conventional administration of the respective bagcontents to a patient.

Accordingly, the triple bag blood collection system of FIG. 1 can beutilized by first obtaining a unit of fresh blood by conventionalvenipuncture with needle 20, and collection of a unit of blood into bag10. Following such collection, donor tube 24 may be sealed by clamping,or by heat sealing in a HEMATRON heat sealing device, sold by the Fenwaldivision of Travenol Laboratories, Inc. Bag 10 contains a small amountof conventional blood cell preservative such as ACD or CPD preservativesolution.

Following the blood collection, and sealing of tube 24, the blood bagsystem is centrifuged, to cause the blood cells in the sample to settleto the bottom of bag 10. After this has been accomplished, cannula 44 ofbag 10 is moved forwardly to rupture diaphragm 42 by manual manipulationof flexible connector tube 17 to push cannula 44 through the diaphragm.The plasma can then be carefully expressed through tube 16 into thesecond bag 12, leaving the packed cells behind in bag 10.

Tube 16 is then usually severed after clamping or heat sealing, and bag10 may be removed from the system, with its packed cells being preservedand stored in a conventional manner until needed for use.

The remaining bags of the system are then placed in a freezingenvironment, for example an ethanol-dry ice bath or mechanicalrefrigeration, to freeze the plasma in bag 12 to the solid state. Afterthe plasma has solidly frozen, the cannula 44 associated with bag 12,and an appropriate portion of flexible connector tube 19, are warmedwith the fingers or the like to permit the cannula to be advanced torupture its associated membrane 42. This opens communication between bag12 and tube 18. Bag 12 is then hung (if desired by means of holes 54) ina refrigerator at a temperature above the plasma freezing point. Thepreferred temperature is approximately 5° C. Bag 14 is placed at aposition vertically below bag 10 to receive cryoprecipitate-poor plasma,as the plasma melts, on a drop by drop basis. The melting plasma in bag12 passes through filtering means 30 and outlet port tube 32, andaccordingly through tubes 18 and 19 into bag 14.

After the plasma has thawed completely and has been filtered into bag14, tube 18 may be sealed in a manner previously described, and severedto permit bag 14 and the cryoprecipitate-poor plasma to be removed forstorage until needed for administration.

An abundant yield of Factor VIII rich cryoprecipitate remains capturedon the filtering means 30 within bag 12. This material may be frozen forstorage if desired.

When the cryoprecipitate is needed for use, it may be dissolved byadding about 10 cc. of sterile physiological saline solution, generallyat room temperature, to the interior of bag 12. This may be done byopening one of the rupturble sheet closures 28 of bag 12 and insertinginto the exposed outlet port 26 a medication injection site of the typecurrently sold by the Fenwal Division of Travelol Laboratories, Inc. Thesaline solution is then added through the medication injection site intothe bag with a syringe and injection needle. The filtering means 30 isthen manipulated and washed with the solution in the bag until all ofthe cryoprecipitate is dissolved in the solution. Thereafter, the bag isinverted so that the outlet ports 26 point downwardly, and the salinesolution, with the dissolved cryoprecipitate, is withdrawn from the bagby means of the syringe needle.

Filtering means 30 is advantageously placed on the side of the bagopposite from the outlet port 26 used in this operation, since the bagmay then be inverted with the outlet port pointed downwardly with thefiltering means 30 out of contact with and above the liquid. Thefiltering means may then be manually squeezed to remove essentially allof the cryoprecipitate solution from it.

That which is claimed is:
 1. In a blood collection system whichcomprises blood compatible container means, including a first, bloodcompatible, sealed container for receiving blood, and a second, bloodcompatible, sealed container, communicating in sterile manner with saidfirst container, for receiving blood plasma from said first container,the improvement comprising a filtering means within saidplasma-receiving container, said filtering means comprising a depthfilter made of open celled foam and being positioned to cover an outletfrom said plasma-receiving container, whereby blood plasma in saidplasma-receiving container may be frozen, and then thawed to form FactorVIII rich cryoprecipitate in said plasma-receiving container, andcryoprecipitate-poor plasma may be transferred from said containerthrough said filtering means and outlet, while said cryoprecipitate isheld in said second container by said filtering means.
 2. The bloodcollection system of claim 1 in which said open cell foam filter is madeof polyurethane.
 3. In a multiple bag blood collection system whichcomprises a plurality of blood-compatible, sealed bags connectedtogether with blood compatible conduit means, and blood collection meanscommunicating with the interior of the first of said bags, theimprovement comprising:filtering means positioned within a second ofsaid bags to cover an outlet from said second bag, said filtering meanscomprising a depth filter made of open celled foam, whereby bloodplasma, expressed from said first bag into said second bag, may befrozen to form Factor VIII rich cryoprecipitate in said second bag, andupon thawing, cryoprecipitate-poor plasma may be transferred from saidsecond bag through said filtering means and outlet, while saidcryoprecipitate is retained in said second bag by said filtering means.4. The blood collection system of claim 3 in which said open cell foamfilter is made of polyurethane.
 5. The blood collection system of claim3 in which said outlet of the second bag is in fluid communication witha third bag, for receiving cryoprecipitate-poor plasma in asepticmanner.